Lezione 19Cromosomi. Territori e domini nucleari

IL CROMOSOMA EUCARIOTICO

Tecniche utilizzate per lo studio della struttura del cromosoma

•Microscopia elettronica•Microscopia a forza ionica•Proteomica•Genomica

Microscopia elettronica e a forza ionica

Zlatanova van Holde capitolo 1

TEM AFM

Imagine di cromosomi umani (AFM)

Zlatanova van Holde capitolo 8

Coesine e condensine

Zlatanova van Holde capitolo 8

Coesine e condensine

Condensine e coesine sono complessi proteici. La coesina forma un anello intorno ai cromatidi fratelli, chevengono separati da una proteasi (separasi)

Centromeri e telomeriIL CROMOSOMA EUCARIOTICO

Centromeri• Principali punti di costrizione del cromosoma • Partecipano all’assemblaggio del cinetocore • I nucleosomi contengono specifiche varianti istoniche,

es. CENPA al posto di H3 • Metilazione H3K9 • Spesso arricchiti in sequenza ripetute e trasposti • Altre proteine:HP-1, CNP-B e CENP-C

https://www.nature.com/articles/35048692

• CENP-B specifically binds to the CENP-B box DNA sequence on the centromere- specific repetitive DNA.

• CENP-C is a critical link between the inner and outer (microtubule-binding) portions of the kinetochore.

• HP1 binds to di- and trimethylated H3K9 (H3K9me2/3) and forms a binding platform for other effecter proteins. Reminiscent to the distribution of cohesin in mitosis, the majority of HP1 proteins in mammals are released from chromosome arms when cells enter mitosis, whereas a small amount of HP1 is retained at the centromere region.

Centromeri - altre proteine

https://www.nature.com/articles/ncb2833?draft=collection

HP1 links centromeric heterochromatin to centromere cohesion in mammals

Choi et al., EMBO reports 2018

Organizzazione 3D di un centromero umano

Organizzazione 3D di un centromero umano

L’avvolgimento della fibrafa sì che CENP-A formiun dominio nella regionerivolta verso il polo. Nellaregione opposta è presenteil normale H3.

Le coesine vengono reclutatedall’eterocromatina. In questaregione più centrale non è chiaro quale sia la struttura.

In questa regione gliistoni sono fortementemetilati a carico dimetilasi specifiche.

TelomeriTelomeres - structure

Interphase- Different types of chromatin:

Active chromatin - Chromatin can become ‘opened’ at gene regulatory regions, which allows RNA polymerase to function.accompanied by ‘active’ covalent modifications of the core histones, including histone H3 methylation on lysine 4 (H3K4me) and H3K9 and H3K27 acetylation At promoters, these modifications can flank a nucleosome-free region immediately upstream of the transcription start site.

‘Low signal state’ chromatin - Chromatin that does not accumulate either active or repressive histone modifications and thus can be considered as unprogrammed.

Actively repressed chromatin - this chromatin is inhibitory to the binding of most transcription factors, including pioneer factors, and thus strongly repress transcription. Such closed domains are typically accompanied by repressive covalent modifications of the core histones, including H3K9 or H3K27 methylation, although usually it does not possess both marks at the same time, thus reflecting the different mechanisms of chromatin repression:

-facultative heterochromatin often forms at developmentally regulated genes, and its level of compaction changes in response to developmental cues and/or environmental signals

-constitutive heterochromatin preferentially assembles at repetitive elements such as satellite DNA and transposons, and maintains high compaction level. Iwafuchi-Doi and Zaret, Development, 2016 Wang et al., Trends in Genetics 2016

Chromosome conformation capture: 3C, 4C, 5C, HI-C

From Babaei et al., Hi-C Chromatin Interaction Networks Predict Co-expression in the Mouse Cortex

Territori cromosomici in un nucleo di cellula umana maschile diploide (colorati con pseudocolori) (modello)

Nucleo di fibroblasto di pollo: specifici fluorofori indicant la presenza di specifici territri cromosomici

Domini associati alla lamina sono trascrizionalmente repressi